Transcription factors of the bHLH and LIM families: synergistic mediators of T cell acute leukemia?
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Adaptor Proteins, Signal Transducing↗
Biomedical subjects
R Baer
Publications and source records attributed to R Baer.
Effect of acetylsalicylic acid on pulmonary gas exchange in patients with severe pneumonia: a pilot study.
BACKGROUND: It has been hypothesized that local release of prostacyclin in acute pneumonia may ablate hypoxic pulmonary vasoconstriction, thus contributing to the impairment of pulmonary gas exchange in these patients. Inhibition of cyclooxygenase pathway could prevent this phenomenon by reducing the release of these metabolites. METHODS: A study was designed to assess the effect of I.V. acetylsalicylic acid (ASA) (2 g) on pulmonary gas exchange in seven patients (age, 64+/-11 [mean+/-SD] years) with unilateral severe pneumonia (PaO2/fraction of inspired oxygen, 168+/-67) needing mechanical ventilation. Respiratory gases, pulmonary and systemic hemodynamics, and ventilation-perfusion (VA/Q) distributions were studied before and 15 and 60 min after the infusion of ASA. RESULTS: At baseline, the amount of shunt (VA/Q ratios <0.005) was 28+/-17% of cardiac output, blood flow to areas with low VA/Q ratios (<0.1, excluding shunt) was 8+/-7%, and the dispersion of pulmonary blood flow distribution (second moment, log SD Q) was 1.45+/-0.49 (normal range, 0.3 to 0.6). Sixty minutes after the infusion of ASA, we observed a mild reduction of the amount of shunt, from 28+/-17% to 23.5+/-13% (p<0.05) without changes in arterial oxygenation. This was associated with a significant increase in mean pulmonary artery pressure (from 21.9+/-3.6 to 24.4+/-5.1 and 23.9+/-5.3 mm Hg, p<0.025 and p=0.1) and pulmonary vascular resistance (from 1.4+/-0.9 to 1.8+/-0.8 and 1.8+/-1.3 mm Hg x min x L(-1) , p<0.002 and p=0.11) 15 and 60 min after ASA, respectively. The ASA plasma levels were within the normal therapeutic range (120+/-7 microg/mL, 15 min, and 113+/-11 microg/mL, 60 min after ASA infusion). CONCLUSIONS: Although there was a modest improvement in intrapulmonary shunt, our results suggest that perfusion of ASA in this small sample of patients with severe pneumonia appears to be of little benefit as complementary treatment for severe hypoxemia.
Adult↗
Influence of dimensionality on deep tunneling rates: A study based on the hydrogen-nickel system.
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Journal Article↗
Mutation of position 52 in ERK2 creates a nonproductive binding mode for adenosine 5'-triphosphate.
Among the protein kinases, an absolutely conserved lysine in subdomain II is required for high catalytic activity. This lysine is known to interact with the substrate ATP, but otherwise its role is not well understood. We have used biochemical and structural methods to investigate the function of this lysine (K52) in phosphoryl transfer reactions catalyzed by the MAP kinase ERK2. The kinetic properties of activated wild-type ERK2 and K52 mutants were examined using the oncoprotein TAL2, myelin basic protein, and a designed synthetic peptide as substrates. The catalytic activities of K52R and K52A ERK2 were lower than that of wild-type ERK2, primarily as a consequence of reductions in kcat. Further, there was little difference in Km for ATP, but the Km,app for peptide substrate was higher for the K52 mutants. The three-dimensional structure of unphosphorylated K52R ERK2 in the absence and presence of bound ATP was determined and compared with the structure of unphosphorylated wild-type ERK2. ATP adopted a well-defined but distinct binding mode in K52R ERK2 compared to the binding mode in the wild-type enzyme. The structural and kinetic data show that mutation of K52 created a nonproductive binding mode for ATP and suggest that K52 is essential for orienting ATP for catalysis.
Adenosine Triphosphate↗
Protein dimerization between Lmo2 (Rbtn2) and Tal1 alters thymocyte development and potentiates T cell tumorigenesis in transgenic mice.
The LMO2 and TAL1 genes were first identified via chromosomal translocations and later found to encode proteins that interact during normal erythroid development. Some T cell leukaemia patients have chromosomal abnormalities involving both genes, implying that LMO2 and TAL1 act synergistically to promote tumorigenesis after their inappropriate co-expression. To test this hypothesis, transgenic mice were made which co-express Lmo2 and Tal1 genes in T cells. Dimers of Lmo2 and Tal1 proteins were formed in thymocytes of double but not single transgenic mice. Furthermore, thymuses of double transgenic mice were almost completely populated by immature T cells from birth, and these mice develop T cell tumours approximately 3 months earlier than those with only the Lmo2 transgene. Thus interaction between these two proteins can alter T cell development and potentiate tumorigenesis. The data also provide formal proof that TAL1 is an oncogene, apparently acting as a tumour promoter in this system.
Adaptor Proteins, Signal Transducing↗
Identification of a RING protein that can interact in vivo with the BRCA1 gene product.
The hereditary breast and ovarian cancer gene, BRCA1, encodes a large polypeptide that contains the cysteine-rich RING motif, a zinc-binding domain found in a variety of regulatory proteins. Here we describe a novel protein that interacts in vivo with the N-terminal region of BRCA1. This BRCA1-associated RING domain (BARD1) protein contains an N-terminal RING motif, three tandem ankyrin repeats, and a C-terminal sequence with significant homology to the phylogenetically conserved BRCT domains that lie near the C terminus of BRCA1. The BARD1/BRCA1 interaction is disrupted by BRCA1 missense mutations that segregate with breast cancer susceptibility, indicating that BARD1 may be involved in mediating tumour suppression by BRCA1.
Amino Acid Sequence↗
Phase space approach for optimizing grid representations: The mapped Fourier method.
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Journal Article↗
Characterization and genomic mapping of a novel leader peptide associated with the human VH4-21 (VH4-34) gene segment.
The human IgVH locus is located on chromosome 14, band q32 and spans approximately 1 mb. Within this locus are approximately 120 VH gene segments that are subdivided into six to seven families based on sequence homology of their coding regions. VH4-21 (VH4-34) is a member of the VH4 family, a family that contains 10 to 15 members. It is expressed in a variety of circumstances including early fetal development, the autoantibody repertoire, and in a highly restricted manner in antibodies that recognize alloantigens on the surface of human red blood cells. Most interesting, however, is the expression of this gene segment in T cells as a semi-germline transcript in conjunction with a nontraditional VH leader peptide. This nonhydrophobic leader sequence, termed "Et" for exon in T cells, has previously been shown to reside within the VH locus. Using YAC and P1 clones, we have identified two copies of this exon, both of which are located in the region of the locus that contains VH4-24 (VH4-34). Characterization of the two exons suggests that they arose by duplication, as flanking DNA is almost identical over a distance of > 5 kb. Preliminary data suggests they are both located > 20 kb upstream of VH4-21 (VH4-34).
Base Sequence↗
Does activation of the TAL1 gene occur in a majority of patients with T-cell acute lymphoblastic leukemia? A pediatric oncology group study.
Almost 25% of patients with T-cell acute lymphoblastic leukemia (T-ALL) have tumor-specific rearrangements of the TAL1 gene. Although TAL1 expression has not been observed in normal lymphocytes, TAL1 gene products are readily detected in leukemic cells that harbor a rearranged TAL1 allele. Hence, it has been proposed that ectopic expression of TAL1 promotes the development of T-ALL. In this report, we show that TAL1 is expressed in the leukemic cells of most patients with T-ALL, including many that do not display an apparent TAL1 gene alteration. A polymorphic dinucleotide repeat in the transcribed sequences of TAL1 was used to determine the allele specificity of TAL1 transcription in primary T-ALL cells. Monoallelic expression of TAL1 was observed in the leukemic cells of all patients (8 of 8) bearing a TAL1 gene rearrangement. In the leukemic cells of patients without detectable TAL1 rearrangements, TAL1 transcription occurred in either a monoallelic (3 of 7 patients) or a biallelic (4 of 7 patients) fashion. Thus, TAL1 activation in these patients may result from subtle alterations in cis-acting regulatory sequences (affecting expression of a single TAL1 allele) or changes in trans-acting factors that control TAL1 transcription (affecting expression of both TAL1 alleles).
Alleles↗
Coordinate expression and developmental role of Id2 protein and TAL1/E2A heterodimer in erythroid progenitor differentiation.
The Id proteins and basic helix-loop-helix (bHLH) proteins play major roles in specifying cell fate decisions in diverse biologic settings. A potential role for Id and TAL1/E2A bHLH genes in hematopoiesis has been suggested by studies on immortalized cell lines. However, it is uncertain whether these observations reflect normal hematopoiesis. We have investigated the expression pattern of Id2 and TAL1/E2A genes in liquid suspension culture of purified hematopoietic progenitor cell (HPCs) undergoing erythroid or granulopoietic differentiation in the first culture week and maturation to terminal cells in the second week. In quiescent, freshly purified HPCs, Id2 mRNA is detected by reverse transcriptase-polymerase chain reaction (RT-PCR), whereas TAL1 and E2A mRNAs are not. At the onset of erythroid differentiation, Id2 mRNA is downregulated, while E2A and TAL1 mRNAs are concomitantly upregulated: their expression is further increased at erythroblast level. Conversely, Id2 is not downmodulated in granulopoietic culture, except for a late decline at day 10 to 12, while TAL1 and E2A are only transiently induced in the first week of granulopoietic differentiation. The expression pattern of the TAL1/E2A heterodimer, as evaluated by mobility shift assay, is consistent with RT-PCR results (except for lower levels of the heterodimer in late erythroid maturation). TAL1 protein level, analyzed by Western blot, shows a pattern consistent with gelshift results. Functional experiments were performed on purified HPCs treated with phosphorothioate antisense oligodeoxynucleotides to Id2 or TAL1 mRNA. The results are strictly consistent with the expression studies: anti-Id2 oligomer (alpha-Id2) causes a significant dose-dependent increase of erythroid colony formation, whereas alpha-TAL1 induces a selective dose-related inhibitory effect on erythroid colonies, as compared with untreated or scrambled oligomer-treated control HPCs. Finally, murine and human glutathione-S-transferase (GST)-Id2 polypeptides compete the TAL1/E2A-specific DNA binding activity when added to the nuclear extracts derived from erythroid culture cells, thus indicating biochemical and suggesting functional interaction of Id2 with the TAL1/E2A complex. These novel observations indicate a coordinate expression and function of an inhibitory Id protein (Id2) and a stimulatory bHLH/bHLH heterodimer (TAL1/E2A) in normal erythroid differentiation.
Adult↗
Specific in vivo association between the bHLH and LIM proteins implicated in human T cell leukemia.
The protein products of proto-oncogenes implicated in T cell acute lymphoblastic leukemia include two distinct families of presumptive transcription factors. RBTN1 and RBTN2 encode highly related proteins that possess cysteine-rich LIM motifs. TAL1, TAL2 and LYL1 encode a unique subgroup of basic helix-loop-helix (bHLH) proteins that share exceptional homology in their bHLH sequences. We have found that RBTN1 and RBTN2 have the ability to interact with each of the leukemogenic bHLH proteins (TAL1, TAL2 and LYL1). These interactions occur in vivo and appear to be mediated by sequences within the LIM and bHLH domains. The LIM-bHLH interactions are highly specific in that RBTN1 and RBTN2 will associate with TAL1, TAL2 and LYL1, but not with other bHLH proteins, including E12, E47, Id1, NHLH1, AP4, MAX, MYC and MyoD1. Moreover, RBTN1 and RBTN2 can interact with TAL1 polypeptides that exist in assembled bHLH heterodimers (e.g. TAL1-E47), suggesting that the RBTN proteins can influence the functional properties of TAL1. Finally, we have identified a subset of leukemia patients that harbor tumor-specific rearrangements of both their RBTN2 and TAL1 genes. Thus, the activated alleles of these genes may promote leukemia cooperatively, perhaps as a result of bHLH-LIM interactions between their protein products.
Amino Acid Sequence↗
The LIM protein RBTN2 and the basic helix-loop-helix protein TAL1 are present in a complex in erythroid cells.
Chromosomal translocations in T-cell acute leukemias can activate genes encoding putative transcription factors such as the LIM proteins RBTN1 and RBTN2 and the DNA-binding basic helix-loop-helix transcription factor TAL1 associated with T-cell acute lymphocytic leukemia. While not expressed in normal T cells, RBTN2 and TAL1 are coexpressed in erythroid cells and are both important for erythroid differentiation. We demonstrate, using anti-RBTN2 and anti-TAL1 antisera, that the LIM protein RBTN2 is not phosphorylated and is complexed with the TAL1 phosphoprotein in the nucleus of erythroid cells. A complex containing both RBTN1 and TAL1 also occurs in a T-cell acute leukemia cell line. Since both RBTN2 and TAL1 are crucial for normal erythropoiesis, these data have important implications for transcription networks therein. Further, since both proteins can be involved in leukemogenesis, these data provide a direct link between proteins activated by chromosomal translocations in T-cell acute leukemia.
Adaptor Proteins, Signal Transducing↗
Positive and negative transcriptional control by the TAL1 helix-loop-helix protein.
Tumor-specific activation of the TAL1 gene is the most common genetic defect associated with T-cell acute lymphoblastic leukemia. The TAL1 gene products possess a basic helix-loop-helix (bHLH) motif, a protein-dimerization and DNA-binding domain found in several transcription factors. TAL1 polypeptides interact, in vitro and in vivo, with class A bHLH proteins (e.g., E47) to form heterodimers with sequence-specific DNA-binding activity. In this study, we show that TAL1 can regulate the transcription of an artificial reporter gene that contains binding sites for bHLH heterodimers involving TAL1. Transcription of the reporter is strongly induced by E47-E47 homodimers and moderately induced by TAL1-E47 heterodimers. Thus, in a cellular environment that allows formation of E47-E47 homodimers (e.g., in the absence of Id regulatory proteins) TAL1 can repress transcription by recruiting E47 into bHLH complexes with less transcriptional activity (i.e., TAL1-E47 heterodimers). However, in other settings TAL1 can activate transcription because TAL1-E47 heterodimers are more resistant to negative regulation by Id proteins. Hence, TAL1 can potentially regulate transcription in either a positive or negative fashion.
Animals↗
Distinguishable patterns of protein-DNA interactions involving complexes of basic helix-loop-helix proteins.
Myogenic factors and TAL1 possess distinguishable DNA binding characteristics when they form a complex with basic helix-loop-helix (bHLH) proteins of class A. These characteristics were evident in electrophoretic mobility shift assays showing that complexes of myogenic factors and HTF4 displayed a relatively high affinity for the enhancer in the muscle creatine kinase gene, whereas TAL1 appeared to greatly attenuate the interaction of HTF4 with this enhancer. In addition, by forming a complex with HTF4 in solution, TAL1 could exert a negative effect on the interactions of HTF4 with elements that include E box motifs of microE2 (CAGCTG) and kappa E2/microE5 (CACCTG) type. Similarly, heterodimers containing TAL1 and the DNA binding domain of E47 exhibited a relatively weak affinity for microE2 and kappa E2/microE5 core motifs. The results of both studies invoked the hypothesis that in vivo TAL1 might act as a negative regulator of microE2 and kappa E2/microE5 sequence motifs by forming a complex with the products of the E2A and HTF4 genes. Support for this hypothesis was obtained by transient expression analyses where TAL1 was found to inhibit the activation effects produced by E2-5 and HTF4a on a reporter gene construct containing repeated microE2 and microE5 motifs, derived from the immunoglobulin gene enhancer.
Amino Acid Sequence↗
Formation of in vivo complexes between the TAL1 and E2A polypeptides of leukemic T cells.
Tumor-specific activation of the TAL1 gene occurs in approximately 25% of patients with T-cell acute lymphoblastic leukemia (T-ALL). The TAL1 gene products possess a basic helix-loop-helix (bHLH) domain that interacts in vitro with the bHLH proteins (E12 and E47) encoded by the E2A locus. We have now applied two independent methods, the two-hybrid procedure and co-immunoprecipitation analysis, to demonstrate that TAL1 and E2A polypeptides also associate in vivo. These studies show that the bHLH domain of TAL1 selectively interacts with the bHLH domains of E12 and E47, but not with the Id1 helix-loop-helix protein. TAL1 does not self-associate to form homodimeric complexes, implying that the in vivo functions of TAL1 depend on heterologous interaction with other bHLH proteins such as E12 and E47. Co-immunoprecipitation analysis revealed the presence of endogenous TAL1/E2A complexes in Jurkat cells, a leukemic line derived from a T-ALL patient. Thus, the malignant properties of TAL1 may be due to obligate interaction with the E2A polypeptides.
Base Sequence↗
HEN1 encodes a 20-kilodalton phosphoprotein that binds an extended E-box motif as a homodimer.
HEN1 and HEN2 encode neuron-specific polypeptides that contain the basic helix-loop-helix (bHLH) motif, a protein dimerization and DNA-binding domain common to several known transcription factors. We now describe characteristics of the HEN1 gene product that are consistent with its postulated role as a transcription factor that functions during development of the mammalian nervous system. Thus, transcription of the HEN1 gene is activated upon the induction of neural differentiation in PC12 cells by nerve growth factor. HEN1 encodes a 20-kDa polypeptide (pp20HEN1) that is phosphorylated exclusively at serine residues and forms dimeric bHLH complexes either by self-association or by heterologous interaction with the E2A gene products (E12 or E47). The resultant HEN1/HEN1 homodimers and HEN1/E2A heterodimers bind DNA in a sequence-specific manner. Moreover, a binding site selection procedure revealed that HEN1-HEN1 homodimers preferentially recognize E-box motifs represented by an 18-bp consensus sequence (GGGNCG CAGCTGCGNCCC). The E-box half-site recognized by HEN1 polypeptides (GGGNCGCAG) is distinct from those of other known bHLH proteins, suggesting that HEN1 binds, an regulates the transcription of, a unique subset of target genes during neural development.
Amino Acid Sequence↗
Preferred sequences for DNA recognition by the TAL1 helix-loop-helix proteins.
Tumor-specific activation of the TAL1 gene is the most common genetic alteration seen in patients with T-cell acute lymphoblastic leukemia. The TAL1 gene products contain the basic helix-loop-helix (bHLH) domain, a protein dimerization and DNA-binding motif common to several known transcription factors. A binding-site selection procedure has now been used to evaluate the DNA recognition properties of TAL1. These studies demonstrate that TAL1 polypeptides do not have intrinsic DNA-binding activity, presumably because of their inability to form bHLH homodimers. However, TAL1 readily interacts with any of the known class A bHLH proteins (E12, E47, E2-2, and HEB) to form heterodimers that bind DNA in a sequence-specific manner. The TAL1 heterodimers preferentially recognize a subset of E-box elements (CANNTG) that can be represented by the consensus sequence AACAGATGGT. This consensus is composed of half-sites for recognition by the participating class A bHLH polypeptide (AACAG) and the TAL1 polypeptide (ATGGT). TAL1 heterodimers with DNA-binding activity are readily detected in nuclear extracts of Jurkat, a leukemic cell line derived from a patient with T-cell acute lymphoblastic leukemia. Hence, TAL1 is likely to bind and regulate the transcription of a unique subset of subordinate target genes, some of which may mediate the malignant function of TAL1 during T-cell leukemogenesis.
Amino Acid Sequence↗
The MAP kinase phosphorylation site of TAL1 occurs within a transcriptional activation domain.
Alteration of the TAL1 gene is the most common genetic lesion found in patients with T cell acute lymphoblastic leukemia. TAL1 encodes a basic helix-loop-helix transcription factor that is phosphorylated on serine residue 122 by the mitogen-activated protein (MAP) kinase ERK1. Here we show that the amino-terminal sequences of TAL1 (residues 1-166) function in vivo as a transcriptional activation domain. Mutation of serine residue 122 reduces the potency of the transactivation domain by more than half. The data suggest that the amino-terminal transactivation domain of TAL1 is positively regulated by S122 phosphorylation and that the functional properties of TAL1 can be influenced by signal transduction pathways that involve the MAP kinases.
Animals↗